1-(3-chlorophenyl)piperazine, common name m-chlorophenylpiperazine (mCPP), is the main metabolite of the widely used antidepressant drug, trazodone, and its less commonly-used analogue nefazodone. It is a serotonin receptor agonist that affects hormone levels, physiology and behaviour, and is known to be a mild hallucinogen with weak ecstasy-like effects. A report by the European Union-funded European Monitoring Centre for Drugs and Drug Addiction (EMCDDA) and Europol describes its increasing abuse across Europe and it is described as the most widely encountered new psychoactive substance since the inception of a European drug monitoring early warning system in 1997 (Europol-EMCDDA Active Monitoring Report on a new psychoactive substance). It is scheduled in several European countries as well as New Zealand, the apparent origin of its use as a “party pill”.
An impediment to testing for mCPP intake is the metabolic production of mCPP by the antidepressant drugs trazodone and nefazodone. Trazodone, systematic name 2-(3-[4-(3-chlorophenyl)piperazin-1-yl]propyl)-[1,2,4]triazolo[4,3-α]pyridin-3(2H)-one, is an antidepressant drug with anxiolytic and hypnotic activity which is metabolized in the liver by hydroxylation, dealkylation and N-oxidation. Staack et al (2007) developed a technique to overcome the possibility of a false-positive test for mCPP intake using gas chromatography and mass spectroscopy (GC-MS). The test rules out trazodone intake by detecting either the parent molecule trazodone or the metabolite hydroxytrazodone. Nefazodone is eliminated as a possible mCPP source by detection of the major metabolites hydroxyl nefazodone and hydroxyethyl deamino hydroxyl nefazodone, both found at higher levels than mCPP. It is reported that mCPP is a minor urinary metabolite of nefazodone in humans, representing less than 1% of total metabolites (Mayol et al. 1994). The drawbacks of the Staack test for mCPP intake are the protracted pre-treatment steps required to prepare derivatives that are amenable to gas chromatography, the expensive and highly-specialised equipment required which is unsuitable and impractical for application outside of the laboratory, and the requirement of an external reference standard of trazodone to confirm the retention time to support an accurate examination of the GC-MS spectra. Furthermore, the requirement of ‘careful screening’ to differentiate between mCPP intake and trazodone implies a method that is non-robust.
Specific binding reactions, such as antibody-antigen interactions, have been used extensively in immunoassays to detect a variety of substances present in tissue extracts. Thus, for example, radioimmunoassays (RIAs) could be used for the determination of mCPP and drugs that produce mCPP as a metabolic product. Radioimmunoassays are very sensitive, but do require radionuclide tracers, for example 125I and 3H. There are no known RIAs for mCPP and drugs that produce mCPP as a metabolic product such as trazodone. Enzyme-linked immunosorbent assays (ELISAs) are a non-radioactive alternative that could be used for the qualitative and quantitative determination of mCPP and drugs that produce mCPP as a metabolic product.
To enable drug screening of mCPP for clinical and forensic toxicology purposes, an economically viable, practical, sensitive and robust test is required. The invention described herein, based on the antibody-antigen interaction, possesses these attributes.